Tumor evolution project

Data used

In this notebook, we are using the tmb_genomic.tsv file generated from the 01-preprocess-data.Rmd script.

Set up

suppressPackageStartupMessages({
  library(tidyverse)
  library(scales)
})

Directories and File Inputs/Outputs

# Detect the \.git\ folder. This will be in the project root directory.
# Use this as the root directory to ensure proper sourcing of functions
# no matter where this is called from.
root_dir <- rprojroot::find_root(rprojroot::has_dir(\.git\))
analysis_dir <- file.path(root_dir, \analyses\, \tmb-vaf-longitudinal\) 
input_dir <- file.path(analysis_dir, \input\)

# File path to results directory
results_dir <-
  file.path(analysis_dir, \results\)
if (!dir.exists(results_dir)) {
  dir.create(results_dir)
}

# Input files
tmb_genomic_file <- file.path(results_dir, \tmb_vaf_genomic.tsv\)
palette_file <- file.path(root_dir, \figures\, \palettes\, \tumor_descriptor_color_palette.tsv\)

# File path to plots directory
plots_dir <-
  file.path(analysis_dir, \plots\)
if (!dir.exists(plots_dir)) {
  dir.create(plots_dir)
}


source(paste0(analysis_dir, \/util/function-create-barplot.R\))
source(paste0(analysis_dir, \/util/function-create-dumbbell-plot.R\))
source(paste0(root_dir, \/figures/scripts/theme.R\))

Read in data and process

# Read and process tmb_genomic file
df_total <- readr::read_tsv(tmb_genomic_file, guess_max = 100000, show_col_types = FALSE) %>% 
  group_by(Kids_First_Participant_ID) %>% 
  mutate(cg_distinct = n_distinct(cancer_group) > 1) # to identify samples with different diagnosis across timepoints

# Are there any samples with both WGS and WXS? 
df_total %>% 
  unique() %>% 
  arrange(Kids_First_Participant_ID, experimental_strategy) %>%
  group_by(Kids_First_Participant_ID) %>%
  dplyr::summarise(experimental_strategy_sum = str_c(experimental_strategy, collapse = \;\)) 
# There are, so let's remove these from downstream analyses.
df <- df_total %>% 
  filter(!experimental_strategy == \WXS\) %>% 
  dplyr::mutate(patient_id = paste(short_histology, Kids_First_Participant_ID, sep = \_\)) %>% 
  distinct(cancer_group, .keep_all = TRUE) %>% 
  summarise(cg_sum = str_c(cancer_group, collapse = \
# A tibble: 37 × 2
   cg_sum                               n
   <chr>                            <int>
 1 High-grade glioma                26116
 2 Meningioma,Medulloblastoma        3713
 3 Atypical Teratoid Rhabdoid Tumor  2944
 4 Rosai-Dorfman disease,Sarcoma     2819
 5 Diffuse midline glioma            2012
 6 Medulloblastoma                   1222
 7 Ependymoma                         906
 8 Low-grade glioma                   737
 9 Chordoma                           477
10 CNS Embryonal tumor                200
# ℹ 27 more rows
# Let's summarize cancer groups with < 10 bs_samples as Other and use this for visualization purposes
cg_sum_df <- df %>% 
  count(cg_sum) %>% 
  dplyr::mutate(cg_sum_n = glue::glue(\{cg_sum} (N={n})\))

df <- df %>% 
  left_join(cg_sum_df, by = c(\cg_sum\)) %>% 
  mutate(cg_plot = case_when(n < 10 ~ \Other\,
                             TRUE ~ cg_sum),
         cg_kids_id = paste(cg_sum, Kids_First_Participant_ID, sep = \_\))

# How many bs_samples per cg_plot?
print(df %>% count(cg_plot) %>% arrange(desc(n))) 
# A tibble: 37 × 2
   cg_plot                              n
   <chr>                            <int>
 1 High-grade glioma                26116
 2 Meningioma,Medulloblastoma        3713
 3 Atypical Teratoid Rhabdoid Tumor  2944
 4 Rosai-Dorfman disease,Sarcoma     2819
 5 Diffuse midline glioma            2012
 6 Medulloblastoma                   1222
 7 Ependymoma                         906
 8 Low-grade glioma                   737
 9 Chordoma                           477
10 CNS Embryonal tumor                200
# ℹ 27 more rows
# Read color palette
palette_df <- readr::read_tsv(palette_file, guess_max = 100000, show_col_types = FALSE)

# Define and order palette
palette <- palette_df$hex_codes
names(palette) <- palette_df$color_names

# length(unique(df$Kids_First_Participant_ID))

TMB per Patient case

We will explore TMB per Kids_First_Participant_ID over time by creating stacked barplots.

# Define parameters for function
ylim = max(df$tmb)
x_value <- df$Kids_First_Participant_ID

# Re-order df
f <- c(\Second Malignancy\, \Unavailable\, \Deceased\, \Recurrence\, \Progressive\, \Diagnosis\) # Level df by timepoints
df_plot <- df %>% 
  dplyr:::mutate(tumor_descriptor = factor(tumor_descriptor),
                 tumor_descriptor = fct_relevel(tumor_descriptor, f)) 

# Run function
fname <- paste0(plots_dir, \/\, \TMB-genomic-total.pdf\)
print(fname)
[1] \/home/rstudio/pbta-tumor-evolution/analyses/tmb-vaf-longitudinal/plots/TMB-genomic-total.pdf\
p <- create_stacked_barplot(tmb_df = df_plot, ylim = ylim, x = x_value, palette = palette)

pdf(file = fname, width = 22, height = 6)
print(p)
dev.off()
png 
  2 

Attention: Hypermutant TMB defined as ≥10 Mb, and Ultrahypermutant TMB defined as ≥100 mutations/Mb in pediatric brain tumors (https://pubmed.ncbi.nlm.nih.gov/29056344/).

Here, we notice that there are samples with high TMB (hyper-mutant samples). Next, we will exclude these samples (threshold >= 10) from downstream analysis. Attention is needed in cases with high number of mutations in only one timepoint as this will lead to un-matched longitudinal samples. We will also remove those so we always have matched longitudinal samples.

# Filter df and remove any samples with single timepoints
df_plot_filter <- df %>%
  filter(!tmb >= 10) %>%
  unique() %>% 
  arrange(Kids_First_Participant_ID, tumor_descriptor) %>%
  group_by(Kids_First_Participant_ID) %>%
  dplyr::summarise(tumor_descriptor_sum = str_c(tumor_descriptor, collapse = \;\)) %>% 
  filter(!tumor_descriptor_sum %in% c(\Diagnosis\, \Progressive\, \Recurrence\, \Second Malignancy\, \Unavailable\, \Deceased\, \Progressive;Progressive\)) %>% 
  dplyr::left_join(df, by = c(\Kids_First_Participant_ID\, \tumor_descriptor_sum\)) %>% 
  mutate(tumor_descriptor = factor(tumor_descriptor),
         tumor_descriptor = fct_relevel(tumor_descriptor, f)) %>% 
  drop_na(tmb) 

# length(unique(df_plot_filter$Kids_First_Participant_ID))

# Define parameters for function
ylim <- max(df_plot_filter$tmb)
df_plot_filter <- df_plot_filter
x_value <- df_plot_filter$cg_kids_id

# Run function
fname <- paste0(plots_dir, \/\, \TMB-genomic-no-hypermutants.pdf\)
print(fname)
[1] \/home/rstudio/pbta-tumor-evolution/analyses/tmb-vaf-longitudinal/plots/TMB-genomic-no-hypermutants.pdf\
p <- create_stacked_barplot(tmb_df = df_plot_filter, ylim = ylim, x = x_value, palette = palette)

pdf(file = fname, width = 25, height = 10)
print(p)
dev.off()
png 
  2 

TMB across timepoints and cancer types per Patient case

We will explore TMB per cancer group over time by creating dumbbell plots. We classified by using cancer types with the highest number of samples (High- and Low-grade gliomas) versus any other cancer groups.

# How many bs_samples per kids_id and cancer group?
# print(table(df_plot_filter$cg_plot))
print(df_plot_filter %>% 
        count(cg_plot, Kids_First_Participant_ID))
# A tibble: 51 × 3
   cg_plot                                Kids_First_Participant_ID     n
   <chr>                                  <chr>                     <int>
 1 Adamantinomatous Craniopharyngioma     PT_YK7AD0KK                  19
 2 Atypical Teratoid Rhabdoid Tumor       PT_DVXE38EX                 198
 3 Atypical Teratoid Rhabdoid Tumor       PT_HE8FBFNA                  16
 4 Atypical Teratoid Rhabdoid Tumor       PT_VTG1S395                  54
 5 CNS Embryonal tumor                    PT_BRVGRXQY                  87
 6 CNS Embryonal tumor,Fibromyxoid lesion PT_98QMQZY7                  68
 7 Diffuse midline glioma                 PT_5NS35B66                  93
 8 Diffuse midline glioma                 PT_JNEV57VK                 456
 9 Diffuse midline glioma                 PT_JSFBMK5V                  94
10 Diffuse midline glioma                 PT_NK8A49X5                 255
# ℹ 41 more rows
# Dumbbell plot per cancer group
cancer_groups <- unique(as.character(df_plot_filter$cg_plot))
cancer_groups <- sort(cancer_groups, decreasing = FALSE)
print(cancer_groups)
 [1] \Adamantinomatous Craniopharyngioma\                  
 [2] \Atypical Teratoid Rhabdoid Tumor\                    
 [3] \CNS Embryonal tumor\                                 
 [4] \CNS Embryonal tumor
for (i in seq_along(cancer_groups)) {
  print(i)
  df_ct_sub <- df_plot_filter %>% 
    filter(cg_plot == cancer_groups [i])
  
      if (i %in% c(3, 7, 8)) {
    print(cancer_groups [i])
    # Define parameters for function
    ylim <- 2
    } else if (i == 2) {
      print(cancer_groups [i])
      # Define parameters for function
      ylim <- 6
      } else {
        print(cancer_groups [i])
        # Define parameters for function
        ylim <- 4
      }
    

    # Name plots
    fname <- paste0(plots_dir, \/\, cancer_groups[i], \-TMB-dumbbell\, \.pdf\)
    print(fname)
    
    # Run function
    p <- create_dumbbell_ct(tmb_df = df_ct_sub, 
                                 ylim = ylim, 
                                 ct_id = cancer_groups[i],
                                 palette = palette)
    pdf(file = fname, width = 12, height = 8)
    print(p)
    dev.off()
}
[1] 1
[1] \Adamantinomatous Craniopharyngioma\
[1] \/home/rstudio/pbta-tumor-evolution/analyses/tmb-vaf-longitudinal/plots/Adamantinomatous Craniopharyngioma-TMB-dumbbell.pdf\

[1] 2
[1] \Atypical Teratoid Rhabdoid Tumor\
[1] \/home/rstudio/pbta-tumor-evolution/analyses/tmb-vaf-longitudinal/plots/Atypical Teratoid Rhabdoid Tumor-TMB-dumbbell.pdf\

[1] 3
[1] \CNS Embryonal tumor\
[1] \/home/rstudio/pbta-tumor-evolution/analyses/tmb-vaf-longitudinal/plots/CNS Embryonal tumor-TMB-dumbbell.pdf\

[1] 4
[1] \CNS Embryonal tumor

[1] 5
[1] \Diffuse midline glioma\
[1] \/home/rstudio/pbta-tumor-evolution/analyses/tmb-vaf-longitudinal/plots/Diffuse midline glioma-TMB-dumbbell.pdf\

[1] 6
[1] \Dysembryoplastic neuroepithelial tumor

[1] 7
[1] \Ependymoma\
[1] \/home/rstudio/pbta-tumor-evolution/analyses/tmb-vaf-longitudinal/plots/Ependymoma-TMB-dumbbell.pdf\

[1] 8
[1] \Ependymoma

[1] 9
[1] \Ganglioglioma\
[1] \/home/rstudio/pbta-tumor-evolution/analyses/tmb-vaf-longitudinal/plots/Ganglioglioma-TMB-dumbbell.pdf\

[1] 10
[1] \Glial-neuronal tumor

[1] 11
[1] \High-grade glioma\
[1] \/home/rstudio/pbta-tumor-evolution/analyses/tmb-vaf-longitudinal/plots/High-grade glioma-TMB-dumbbell.pdf\

[1] 12
[1] \Low-grade glioma\
[1] \/home/rstudio/pbta-tumor-evolution/analyses/tmb-vaf-longitudinal/plots/Low-grade glioma-TMB-dumbbell.pdf\

[1] 13
[1] \Low-grade glioma

[1] 14
[1] \Low-grade glioma

[1] 15
[1] \Medulloblastoma\
[1] \/home/rstudio/pbta-tumor-evolution/analyses/tmb-vaf-longitudinal/plots/Medulloblastoma-TMB-dumbbell.pdf\

[1] 16
[1] \Medulloblastoma

[1] 17
[1] \Meningioma\
[1] \/home/rstudio/pbta-tumor-evolution/analyses/tmb-vaf-longitudinal/plots/Meningioma-TMB-dumbbell.pdf\

[1] 18
[1] \Neuroblastoma

[1] 19
[1] \Pilocytic astrocytoma

[1] 20
[1] \Schwannoma\
[1] \/home/rstudio/pbta-tumor-evolution/analyses/tmb-vaf-longitudinal/plots/Schwannoma-TMB-dumbbell.pdf\

Total number of mutations across timepoints and biospecimen sample per Patient case

Here, we want to explore the number of mutations per timepoint and biospecimen sample per patient case.

samples <- unique(as.character(df_plot_filter$Kids_First_Participant_ID))
print(samples)
 [1] \PT_02J5CWN5\ \PT_0DWRY9ZX\ \PT_1ZAWNGWT\ \PT_2FVTD0WR\ \PT_2MZPGZN1\
 [6] \PT_2YT37G8P\ \PT_37B5JRP1\ \PT_39H4JN6H\ \PT_3GYW6P6P\ \PT_3P3HARZ2\
[11] \PT_3R0P995B\ \PT_3T3VGWC6\ \PT_3VCS1PPF\ \PT_5NS35B66\ \PT_5ZPPR06P\
[16] \PT_62G82T6Q\ \PT_82MX6J77\ \PT_89XRZBSG\ \PT_962TCBVR\ \PT_98QMQZY7\
[21] \PT_99S5BPE3\ \PT_B5DQ8FF0\ \PT_BRVGRXQY\ \PT_BZCJMEX8\ \PT_CWXSP19D\
[26] \PT_CXT81GRM\ \PT_DFQAH7RS\ \PT_DNAJYFZT\ \PT_DVXE38EX\ \PT_FN4GEEFR\
[31] \PT_HE8FBFNA\ \PT_HHG37M6W\ \PT_JNEV57VK\ \PT_JSFBMK5V\ \PT_KMHGNCNR\
[36] \PT_NK8A49X5\ \PT_P571HTNK\ \PT_PF04R0BH\ \PT_PR4YBBH3\ \PT_QH9H491G\
[41] \PT_S2SQJVGK\ \PT_T4VN7ZRB\ \PT_TKWTTRQ7\ \PT_TP6GS00H\ \PT_TRZ1N1HQ\
[46] \PT_VTG1S395\ \PT_XA98HG1C\ \PT_XHYBZKCX\ \PT_YK7AD0KK\ \PT_Z4GS3ZQQ\
[51] \PT_ZMKMKCFQ\
for (i in seq_along(samples)) {
  print(i)
  tmb_sub <- df_plot_filter %>%
    filter(Kids_First_Participant_ID == samples[i])
  
  # Define parameters for function
  ylim = max(df_plot_filter$mutation_count)
 
  # Run function
  p <- create_barplot_sample(tmb_df = tmb_sub,
                             ylim = ylim,
                             sid = samples[i],
                             palette = palette)
  print(p)
}
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sessionInfo()
R version 4.2.3 (2023-03-15)
Platform: x86_64-pc-linux-gnu (64-bit)
Running under: Ubuntu 22.04.2 LTS

Matrix products: default
BLAS:   /usr/lib/x86_64-linux-gnu/openblas-pthread/libblas.so.3
LAPACK: /usr/lib/x86_64-linux-gnu/openblas-pthread/libopenblasp-r0.3.20.so

locale:
 [1] LC_CTYPE=en_US.UTF-8       LC_NUMERIC=C              
 [3] LC_TIME=en_US.UTF-8        LC_COLLATE=en_US.UTF-8    
 [5] LC_MONETARY=en_US.UTF-8    LC_MESSAGES=en_US.UTF-8   
 [7] LC_PAPER=en_US.UTF-8       LC_NAME=C                 
 [9] LC_ADDRESS=C               LC_TELEPHONE=C            
[11] LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C       

attached base packages:
[1] grid      stats     graphics  grDevices utils     datasets  methods  
[8] base     

other attached packages:
 [1] ggthemes_4.2.4  scales_1.2.1    lubridate_1.9.2 forcats_1.0.0  
 [5] stringr_1.5.0   dplyr_1.1.1     purrr_1.0.1     readr_2.1.4    
 [9] tidyr_1.3.0     tibble_3.2.1    ggplot2_3.4.0   tidyverse_2.0.0

loaded via a namespace (and not attached):
 [1] highr_0.10       bslib_0.4.2      compiler_4.2.3   pillar_1.9.0    
 [5] jquerylib_0.1.4  tools_4.2.3      bit_4.0.5        digest_0.6.31   
 [9] timechange_0.2.0 jsonlite_1.8.4   evaluate_0.20    lifecycle_1.0.3 
[13] gtable_0.3.3     pkgconfig_2.0.3  rlang_1.1.0      cli_3.6.1       
[17] parallel_4.2.3   yaml_2.3.7       xfun_0.38        fastmap_1.1.1   
[21] withr_2.5.0      knitr_1.42       generics_0.1.3   vctrs_0.6.2     
[25] sass_0.4.5       hms_1.1.3        bit64_4.0.5      rprojroot_2.0.3 
[29] tidyselect_1.2.0 glue_1.6.2       R6_2.5.1         fansi_1.0.4     
[33] vroom_1.6.1      rmarkdown_2.21   farver_2.1.1     tzdb_0.3.0      
[37] magrittr_2.0.3   htmltools_0.5.5  colorspace_2.1-0 labeling_0.4.2  
[41] utf8_1.2.3       stringi_1.7.12   munsell_0.5.0    cachem_1.0.7    
[45] crayon_1.5.2    
---
title: "Explore TMB and number of mutations across multiple timepoints of the PBTA Cohort"
author: "Antonia Chroni <chronia@chop.edu> for D3B"
date: "2023"
output:
  html_notebook:
    toc: TRUE
    toc_float: TRUE
---

#### Tumor evolution project 

### Data used 
In this notebook, we are using the `tmb_genomic.tsv` file generated from the `01-preprocess-data.Rmd` script.

# Set up
```{r load-library}
suppressPackageStartupMessages({
  library(tidyverse)
  library(scales)
})
```

# Directories and File Inputs/Outputs
```{r set-dir-and-file-names}
# Detect the ".git" folder. This will be in the project root directory.
# Use this as the root directory to ensure proper sourcing of functions
# no matter where this is called from.
root_dir <- rprojroot::find_root(rprojroot::has_dir(".git"))
scratch_dir <- file.path(root_dir, "scratch")
analysis_dir <- file.path(root_dir, "analyses", "tmb-vaf-longitudinal") 
input_dir <- file.path(analysis_dir, "input")

# File path to results directory
results_dir <-
  file.path(analysis_dir, "results")
if (!dir.exists(results_dir)) {
  dir.create(results_dir)
}

# Input files
tmb_genomic_file <- file.path(scratch_dir, "tmb_vaf_genomic.tsv")
palette_file <- file.path(root_dir, "figures", "palettes", "tumor_descriptor_color_palette.tsv")

# File path to plots directory
plots_dir <-
  file.path(analysis_dir, "plots")
if (!dir.exists(plots_dir)) {
  dir.create(plots_dir)
}

# File path to dumbbell plots directory
dumbbell_plots_dir <-
  file.path(analysis_dir, "plots", "dumbbell")
if (!dir.exists(dumbbell_plots_dir )) {
  dir.create(dumbbell_plots_dir )
}



source(paste0(analysis_dir, "/util/function-create-barplot.R"))
source(paste0(analysis_dir, "/util/function-create-dumbbell-plot.R"))
source(paste0(root_dir, "/figures/scripts/theme.R"))
```

# Read in data and process
```{r read_input_files}
# Read and process tmb_genomic file
df_total <- readr::read_tsv(tmb_genomic_file, guess_max = 100000, show_col_types = FALSE) %>% 
  group_by(Kids_First_Participant_ID) %>% 
  mutate(cg_distinct = n_distinct(cancer_group) > 1) # to identify samples with different diagnosis across timepoints

# Are there any samples with both WGS and WXS? 
df_total %>% 
  unique() %>% 
  arrange(Kids_First_Participant_ID, experimental_strategy) %>%
  group_by(Kids_First_Participant_ID) %>%
  dplyr::summarise(experimental_strategy_sum = str_c(experimental_strategy, collapse = ";")) 

# There are, so let's remove these from downstream analyses.
df <- df_total %>% 
  filter(!experimental_strategy == "WXS") %>% 
  dplyr::mutate(patient_id = paste(short_histology, Kids_First_Participant_ID, sep = "_")) %>% 
  distinct(cancer_group, .keep_all = TRUE) %>% 
  summarise(cg_sum = str_c(cancer_group, collapse = ",")) %>% # to identify cases with multiple diagnosis
  left_join(df_total, by = c("Kids_First_Participant_ID")) %>% 
  select(Kids_First_Participant_ID, Kids_First_Biospecimen_ID, cg_sum, cancer_group, short_histology, tumor_descriptor, descriptors, tumor_descriptor_sum, tmb, mutation_count)

# How many bs_samples per cg_sum?
print(df %>% count(cg_sum) %>% arrange(desc(n))) 

# Let's summarize cancer groups with < 10 bs_samples as Other and use this for visualization purposes
cg_sum_df <- df %>% 
  count(cg_sum) %>% 
  dplyr::mutate(cg_sum_n = glue::glue("{cg_sum} (N={n})"))

df <- df %>% 
  left_join(cg_sum_df, by = c("cg_sum")) %>% 
  mutate(cg_plot = case_when(n < 10 ~ "Other",
                             TRUE ~ cg_sum),
         cg_kids_id = paste(cg_sum, Kids_First_Participant_ID, sep = "_"))

# How many bs_samples per cg_plot?
print(df %>% count(cg_plot) %>% arrange(desc(n))) 

# Read color palette
palette_df <- readr::read_tsv(palette_file, guess_max = 100000, show_col_types = FALSE)

# Define and order palette
palette <- palette_df$hex_codes
names(palette) <- palette_df$color_names

# length(unique(df$Kids_First_Participant_ID))
```

# TMB per Patient case
We will explore TMB per `Kids_First_Participant_ID` over time by creating stacked barplots.

```{r create-stacked-barplot, fig.width = 22, fig.height = 6, fig.fullwidth = TRUE}
# Define parameters for function
ylim = max(df$tmb)
x_value <- df$Kids_First_Participant_ID

# Re-order df
f <- c("Second Malignancy", "Unavailable", "Deceased", "Recurrence", "Progressive", "Diagnosis") # Level df by timepoints
df_plot <- df %>% 
  dplyr:::mutate(tumor_descriptor = factor(tumor_descriptor),
                 tumor_descriptor = fct_relevel(tumor_descriptor, f)) 

# Run function
fname <- paste0(plots_dir, "/", "TMB-genomic-total.pdf")
print(fname)
p <- create_stacked_barplot(tmb_df = df_plot, ylim = ylim, x = x_value, palette = palette)
pdf(file = fname, width = 22, height = 6)
print(p)
dev.off()
```
Attention: Hypermutant TMB defined as ≥10 Mb, and Ultrahypermutant TMB defined as ≥100 mutations/Mb in pediatric brain tumors (https://pubmed.ncbi.nlm.nih.gov/29056344/).

Here, we notice that there are samples with high TMB (hyper-mutant samples). Next, we will exclude these samples (threshold >= 10) from downstream analysis. Attention is needed in cases with high number of mutations in only one timepoint as this will lead to un-matched longitudinal samples. We will also remove those so we always have matched longitudinal samples.

```{r create-stacked-barplot-filter, fig.width = 25, fig.height = 10, fig.fullwidth = TRUE}
# Filter df and remove any samples with single timepoints
df_plot_filter <- df %>%
  filter(!tmb >= 10) %>%
  unique() %>% 
  arrange(Kids_First_Participant_ID, tumor_descriptor) %>%
  group_by(Kids_First_Participant_ID) %>%
  dplyr::summarise(tumor_descriptor_sum = str_c(tumor_descriptor, collapse = ";")) %>% 
  filter(!tumor_descriptor_sum %in% c("Diagnosis", "Progressive", "Recurrence", "Second Malignancy", "Unavailable", "Deceased", "Progressive;Progressive")) %>% 
  dplyr::left_join(df, by = c("Kids_First_Participant_ID", "tumor_descriptor_sum")) %>% 
  mutate(tumor_descriptor = factor(tumor_descriptor),
         tumor_descriptor = fct_relevel(tumor_descriptor, f)) %>% 
  drop_na(tmb) 

# length(unique(df_plot_filter$Kids_First_Participant_ID))

# Define parameters for function
ylim <- max(df_plot_filter$tmb)
df_plot_filter <- df_plot_filter
x_value <- df_plot_filter$cg_kids_id

# Run function
fname <- paste0(plots_dir, "/", "TMB-genomic-no-hypermutants.pdf")
print(fname)
p <- create_stacked_barplot(tmb_df = df_plot_filter, ylim = ylim, x = x_value, palette = palette)
pdf(file = fname, width = 25, height = 10)
print(p)
dev.off()
```

# TMB across timepoints and cancer types per Patient case
We will explore TMB per cancer group over time by creating dumbbell plots. We classified by using cancer types with the highest number of samples (High- and Low-grade gliomas) versus any other cancer groups.

```{r create-dumbbell-ct, fig.width = 12, fig.height = 8, fig.fullwidth = TRUE}
# How many bs_samples per kids_id and cancer group?
# print(table(df_plot_filter$cg_plot))
print(df_plot_filter %>% 
        count(cg_plot, Kids_First_Participant_ID))

# Dumbbell plot per cancer group
cancer_groups <- unique(as.character(df_plot_filter$cg_plot))
cancer_groups <- sort(cancer_groups, decreasing = FALSE)
print(cancer_groups)

for (i in seq_along(cancer_groups)) {
  print(i)
  df_ct_sub <- df_plot_filter %>% 
    filter(cg_plot == cancer_groups [i])
  
      if (i %in% c(3, 7, 8)) {
    print(cancer_groups [i])
    # Define parameters for function
    ylim <- 2
    } else if (i == 2) {
      print(cancer_groups [i])
      # Define parameters for function
      ylim <- 6
      } else {
        print(cancer_groups [i])
        # Define parameters for function
        ylim <- 4
      }
    

    # Name plots
    fname <- paste0(dumbbell_plots_dir, "/", cancer_groups[i], "-TMB-dumbbell", ".pdf")
    print(fname)
    
    # Run function
    p <- create_dumbbell_ct(tmb_df = df_ct_sub, 
                                 ylim = ylim, 
                                 ct_id = cancer_groups[i],
                                 palette = palette)
    pdf(file = fname, width = 12, height = 8)
    print(p)
    dev.off()
}
```

# Total number of mutations across timepoints and biospecimen sample per Patient case
Here, we want to explore the number of mutations per timepoint and biospecimen sample per patient case.

```{r create-barplot-sample, fig.width = 5, fig.height = 4, fig.fullwidth = TRUE}
samples <- unique(as.character(df_plot_filter$Kids_First_Participant_ID))
print(samples)

for (i in seq_along(samples)) {
  print(i)
  tmb_sub <- df_plot_filter %>%
    filter(Kids_First_Participant_ID == samples[i])
  
  # Define parameters for function
  ylim = max(df_plot_filter$mutation_count)
 
  # Run function
  p <- create_barplot_sample(tmb_df = tmb_sub,
                             ylim = ylim,
                             sid = samples[i],
                             palette = palette)
  print(p)
}
```

```{r echo=TRUE}
sessionInfo()
```
